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  1 / 1046 MEDLINE  
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[PMID]:27775150
[Au] Autor:Schmölzer K; Lemmerer M; Gutmann A; Nidetzky B
[Ad] Endereço:Austrian Centre of Industrial Biotechnology, Petersgasse 14, 8010, Graz, Austria.
[Ti] Título:Integrated process design for biocatalytic synthesis by a Leloir Glycosyltransferase: UDP-glucose production with sucrose synthase.
[So] Source:Biotechnol Bioeng;114(4):924-928, 2017 04.
[Is] ISSN:1097-0290
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Nucleotide sugar-dependent ("Leloir") glycosyltransferases (GTs), represent a new paradigm for the application of biocatalytic glycosylations to the production of fine chemicals. However, it remains to be shown that GT processes meet the high efficiency targets of industrial biotransformations. We demonstrate in this study of uridine-5'-diphosphate glucose (UDP-glc) production by sucrose synthase (from Acidithiobacillus caldus) that a holistic process design, involving coordinated development of biocatalyst production, biotransformation, and downstream processing (DSP) was vital for target achievement at ∼100 g scale synthesis. Constitutive expression in Escherichia coli shifted the recombinant protein production mainly to the stationary phase and enhanced the specific enzyme activity to a level (∼480 U/g ) suitable for whole-cell biotransformation. The UDP-glc production had excellent performance metrics of ∼100 g /L, 86% yield (based on UDP), and a total turnover number of 103 g /g at a space-time yield of 10 g/L/h. Using efficient chromatography-free DSP, the UDP-glc was isolated in a single batch with ≥90% purity and in 73% isolated yield. Overall, the process would allow production of ∼0.7 kg of isolated product/L E. coli bioreactor culture, thus demonstrating how integrated process design promotes the practical use of a GT conversion. Biotechnol. Bioeng. 2017;114: 924-928. © 2016 Wiley Periodicals, Inc.
[Mh] Termos MeSH primário: Glucosiltransferases/metabolismo
Glicosiltransferases/metabolismo
Uridina Difosfato Glucose/análise
Uridina Difosfato Glucose/metabolismo
[Mh] Termos MeSH secundário: Proteínas de Bactérias/metabolismo
Escherichia coli/metabolismo
Glicosilação
Nucleotídeos
Proteínas Recombinantes/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (Nucleotides); 0 (Recombinant Proteins); EC 2.4.- (Glycosyltransferases); EC 2.4.1.- (Glucosyltransferases); EC 2.4.1.13 (sucrose synthase); V50K1D7P4Y (Uridine Diphosphate Glucose)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171216
[Lr] Data última revisão:
171216
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:161025
[St] Status:MEDLINE
[do] DOI:10.1002/bit.26204


  2 / 1046 MEDLINE  
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[PMID]:28741460
[Au] Autor:Wei S; Zhang XY; Sun Y; Conway LP; Liu L
[Ad] Endereço:Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing. China.
[Ti] Título:Discovery and Biochemical Characterization of UDP-Glucose Dehydrogenase from Akkermansia muciniphila.
[So] Source:Protein Pept Lett;24(8):735-741, 2017.
[Is] ISSN:1875-5305
[Cp] País de publicação:Netherlands
[La] Idioma:eng
[Ab] Resumo:BACKGROUND: The biocatalytic oxidation of UDP-glucose in the presence of NAD+ is catalyzed by UDP-glucose dehydrogenases. OBJECTIVES: The main objective of this study was the characterization of a UDP-glucose dehydrogenase (AmUGD) from Akkermansia muciniphila, a bacterium originally isolated from human faeces in an anaerobic medium containing gastric mucin as the sole carbon source. METHODS: The biochemical analysis of AmUGD was performed using a plate reader-based assay measuring the reaction by-product NADH. Furthermore, HPLC- and MALDI-ToF-MS- based methods were used for the enzyme characterization. RESULTS: The recombinant form of the protein was expressed in E. coli and the purified enzyme exhibited optimum levels of activity at 37°C and pH 9.0. While the enzyme is active in the absence of metal ions, the presence of Zn2+ ions results in markedly enhanced levels of catalysis. CONCLUSION: This study describes the first characterization of a nucleotide-processing enzyme from A. muciniphila. The ease of expression and purification of this enzyme make it ideal for biotechnological applications such as the enzymatic synthesis of nucleotide sugars, which may in turn be used for the synthesis of complex carbohydrates or glycoconjugates.
[Mh] Termos MeSH primário: Proteínas de Bactérias/metabolismo
NAD/metabolismo
Uridina Difosfato Glucose Desidrogenase/metabolismo
Uridina Difosfato Glucose/metabolismo
Verrucomicrobia/química
[Mh] Termos MeSH secundário: Proteínas de Bactérias/genética
Cátions Bivalentes
Clonagem Molecular
Escherichia coli/genética
Escherichia coli/metabolismo
Expressão Gênica
Temperatura Alta
Concentração de Íons de Hidrogênio
Cinética
NAD/química
Plasmídeos/química
Plasmídeos/metabolismo
Proteínas Recombinantes/genética
Proteínas Recombinantes/metabolismo
Especificidade por Substrato
Uridina Difosfato Glucose/química
Uridina Difosfato Glucose Desidrogenase/genética
Verrucomicrobia/enzimologia
Zinco/química
Zinco/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (Cations, Divalent); 0 (Recombinant Proteins); 0U46U6E8UK (NAD); EC 1.1.1.22 (Uridine Diphosphate Glucose Dehydrogenase); J41CSQ7QDS (Zinc); V50K1D7P4Y (Uridine Diphosphate Glucose)
[Em] Mês de entrada:1711
[Cu] Atualização por classe:171128
[Lr] Data última revisão:
171128
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170726
[St] Status:MEDLINE
[do] DOI:10.2174/0929866524666170724111147


  3 / 1046 MEDLINE  
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[PMID]:28741459
[Au] Autor:Li Q; Huang YY; Conway LP; He M; Wei S; Huang K; Duan XC; Flitsch SL; Voglmeir J
[Ad] Endereço:Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing. China.
[Ti] Título:Discovery and Biochemical Characterization of a Thermostable Glucose-1-phosphate Nucleotidyltransferase from Thermodesulfatator indicus.
[So] Source:Protein Pept Lett;24(8):729-734, 2017.
[Is] ISSN:1875-5305
[Cp] País de publicação:Netherlands
[La] Idioma:eng
[Ab] Resumo:BACKGROUND: The biosynthesis of NDP-glucoses is based on the nucleotide transfer from NTP donor substrates to glucose-1-phosphates catalyzed by glucose-1-phosphate nucleotidyltransferases. OBJECTIVES: The cloning and biochemical characterization of a glucose-1-phosphate nucleotidyltransferase (TiGPNT) from the deep sea bacterium Thermodesulfatator indicus. METHODS: The biochemical parameters of recombinant TiGPNT were determined using a plate reader-based coupled enzymatic assay, in which the reaction product UDP-glucose is oxidized in the presence of NAD+ forming UDP-Glucuronic acid and NADH. The substrate promiscuity of the enzyme was determined using thin-layer chromatography and MALDI-ToF mass spectrometry. RESULTS: TiGPNT was recombinantly expressed under the control of the T7 promoter in Escherichia coli and could be successfully enriched by heat treatment at 80°C for 30 min. The obtained enzyme worked best at pH 7.5 and the optimum reaction temperature was determined to be 50°C. Interestingly, TiGPNT could fully retain its activity even after extended incubation periods at temperatures of up to 80°C. The enzyme was strongly inhibited in the presence of Cu2+ and Fe2+ ions and EDTA. Among the tested glycosyl donor substrates, TiGPNT showed strict specificity towards glucose-1-phosphate. At the same time, TiGPNT was highly promiscuous towards all tested nucleotide donor substrates. CONCLUSION: TiGPNT shows comparable biochemical features in regards to pH optima, temperature optima and the substrate specificity to characterized glucose-1-phosphate nucleotidyltransferase from other species. The enzyme was capable of utilizing glucose-1-phosphate and all tested nucleoside triphosphate donors as substrates. The high activity of the enzyme and the simple purification protocol make TiGPNT an interesting new biocatalyst for the synthesis of glucose-diphospho nucleosides.
[Mh] Termos MeSH primário: Bactérias/química
Proteínas de Bactérias/metabolismo
Glucofosfatos/química
NAD/química
Uridina Difosfato Glucose/química
[Mh] Termos MeSH secundário: Organismos Aquáticos
Bactérias/enzimologia
Proteínas de Bactérias/genética
Clonagem Molecular
Estabilidade Enzimática
Escherichia coli/genética
Escherichia coli/metabolismo
Expressão Gênica
Glucofosfatos/metabolismo
Temperatura Alta
Concentração de Íons de Hidrogênio
Cinética
NAD/metabolismo
Nucleotidiltransferases/genética
Nucleotidiltransferases/metabolismo
Plasmídeos/química
Plasmídeos/metabolismo
Proteínas Recombinantes/genética
Proteínas Recombinantes/metabolismo
Especificidade por Substrato
Uridina Difosfato Glucose/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (Glucosephosphates); 0 (Recombinant Proteins); 0U46U6E8UK (NAD); CIX3U01VAU (glucose-1-phosphate); EC 2.7.7.- (Nucleotidyltransferases); V50K1D7P4Y (Uridine Diphosphate Glucose)
[Em] Mês de entrada:1711
[Cu] Atualização por classe:171128
[Lr] Data última revisão:
171128
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170726
[St] Status:MEDLINE
[do] DOI:10.2174/0929866524666170724110408


  4 / 1046 MEDLINE  
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[PMID]:28752893
[Au] Autor:Honarmand Ebrahimi K; Carr SB; McCullagh J; Wickens J; Rees NH; Cantley J; Armstrong FA
[Ad] Endereço:Department Chemistry, University of Oxford, UK.
[Ti] Título:The radical-SAM enzyme Viperin catalyzes reductive addition of a 5'-deoxyadenosyl radical to UDP-glucose in vitro.
[So] Source:FEBS Lett;591(16):2394-2405, 2017 Aug.
[Is] ISSN:1873-3468
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Viperin, a radical-S-adenosylmethionine (SAM) enzyme conserved from fungi to humans, can restrict replication of many viruses. Neither the molecular mechanism underlying the antiviral activity of Viperin, nor its exact physiological function, is understood: most importantly, no radical-SAM activity has been discovered for Viperin. Here, using electron paramagnetic resonance (EPR) spectroscopy, mass spectrometry, and NMR spectroscopy, we show that uridine diphosphate glucose (UDP-glucose) is a substrate of a fungal Viperin (58% pairwise identity with human Viperin at the amino acid level) in vitro. Structural homology modeling and docking experiments reveal a highly conserved binding pocket in which the position of UDP-glucose is consistent with our experimental data regarding catalytic addition of a 5'-deoxyadenosyl radical and a hydrogen atom to UDP-glucose.
[Mh] Termos MeSH primário: Biocatálise
Desoxiadenosinas/metabolismo
Proteínas Fúngicas/metabolismo
S-Adenosilmetionina/metabolismo
Uridina Difosfato Glucose/metabolismo
[Mh] Termos MeSH secundário: Sequência de Aminoácidos
Sequência Conservada
Desoxiadenosinas/química
Radicais Livres/química
Radicais Livres/metabolismo
Proteínas Fúngicas/química
Hidrogênio
Simulação de Acoplamento Molecular
Oxirredução
Conformação Proteica
Sordariales/enzimologia
[Pt] Tipo de publicação:LETTER
[Nm] Nome de substância:
0 (Deoxyadenosines); 0 (Free Radicals); 0 (Fungal Proteins); 7LP2MPO46S (S-Adenosylmethionine); 7YNJ3PO35Z (Hydrogen); V50K1D7P4Y (Uridine Diphosphate Glucose)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171002
[Lr] Data última revisão:
171002
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170729
[St] Status:MEDLINE
[do] DOI:10.1002/1873-3468.12769


  5 / 1046 MEDLINE  
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[PMID]:28739255
[Au] Autor:Tsurumaki H; Katano H; Sato K; Imai R; Niino S; Hirabayashi Y; Ichikawa S
[Ad] Endereço:Laboratory for Animal Cell Engineering, Niigata University of Pharmacy and Applied Life Sciences (NUPALS), 265-1 Higashijima, Akiha-ku, Niigata-shi, Niigata 956-8603, Japan.
[Ti] Título:WP1066, a small molecule inhibitor of the JAK/STAT3 pathway, inhibits ceramide glucosyltransferase activity.
[So] Source:Biochem Biophys Res Commun;491(2):265-270, 2017 Sep 16.
[Is] ISSN:1090-2104
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:WP1066 is a well-known inhibitor of the JAK/STAT3 signaling pathway. By a screen of known small molecule inhibitors of various enzymes and protein factors, we identified WP1066 as a ceramide glucosyltransferase inhibitor. Ceramide glucosyltransferase catalyzes the first glycosylation step during glycosphingolipid synthesis. We found that WP1066 inhibited the activity of ceramide glucosyltransferase with an IC of 7.2 µM, and that its action was independent of JAK/STAT3 pathway blockade. Moreover, the modes of inhibition of ceramide glucosyltransferase were uncompetitive with respect to both C -NBD-cermide and UDP-glucose.
[Mh] Termos MeSH primário: Inibidores Enzimáticos/farmacologia
Glucosiltransferases/antagonistas & inibidores
Melanócitos/efeitos dos fármacos
Piridinas/farmacologia
Bibliotecas de Moléculas Pequenas/farmacologia
Tirfostinas/farmacologia
[Mh] Termos MeSH secundário: 4-Cloro-7-nitrobenzofurazano/análogos & derivados
4-Cloro-7-nitrobenzofurazano/química
4-Cloro-7-nitrobenzofurazano/metabolismo
Animais
Linhagem Celular
Ceramidas/química
Ceramidas/metabolismo
Ensaios Enzimáticos
Inibidores Enzimáticos/química
Expressão Gênica
Glucosiltransferases/genética
Glucosiltransferases/metabolismo
Seres Humanos
Janus Quinases/antagonistas & inibidores
Janus Quinases/genética
Janus Quinases/metabolismo
Cinética
Melanócitos/citologia
Melanócitos/enzimologia
Piridinas/química
Ratos
Fator de Transcrição STAT3/antagonistas & inibidores
Fator de Transcrição STAT3/genética
Fator de Transcrição STAT3/metabolismo
Transdução de Sinais
Bibliotecas de Moléculas Pequenas/química
Tirfostinas/química
Uridina Difosfato Glucose/química
Uridina Difosfato Glucose/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Ceramides); 0 (Enzyme Inhibitors); 0 (Pyridines); 0 (STAT3 Transcription Factor); 0 (STAT3 protein, human); 0 (Small Molecule Libraries); 0 (Tyrphostins); 0 (WP1066); 86701-10-2 (N-(7-(4-nitrobenzo-2-oxa-1,3-diazole))-6-aminocaproyl sphingosine); EC 2.4.1.- (Glucosyltransferases); EC 2.4.1.80 (ceramide glucosyltransferase); EC 2.7.10.2 (Janus Kinases); EQF2794IRE (4-Chloro-7-nitrobenzofurazan); V50K1D7P4Y (Uridine Diphosphate Glucose)
[Em] Mês de entrada:1708
[Cu] Atualização por classe:170824
[Lr] Data última revisão:
170824
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170726
[St] Status:MEDLINE


  6 / 1046 MEDLINE  
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[PMID]:28531287
[Au] Autor:Julius C; Yuzenkova Y
[Ad] Endereço:Centre for Bacterial Cell Biology, Institute for Cell and Molecular Biosciences, Newcastle University, Baddiley-Clark Building, Richardson Road, Newcastle upon Tyne, NE2 4AX, UK.
[Ti] Título:Bacterial RNA polymerase caps RNA with various cofactors and cell wall precursors.
[So] Source:Nucleic Acids Res;45(14):8282-8290, 2017 Aug 21.
[Is] ISSN:1362-4962
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Bacterial RNA polymerase is able to initiate transcription with adenosine-containing cofactor NAD+, which was proposed to result in a portion of cellular RNAs being 'capped' at the 5' end with NAD+, reminiscent of eukaryotic cap. Here we show that, apart from NAD+, another adenosine-containing cofactor FAD and highly abundant uridine-containing cell wall precursors, UDP-Glucose and UDP-N-acetylglucosamine are efficiently used to initiate transcription in vitro. We show that the affinity to NAD+ and UDP-containing factors during initiation is much lower than their cellular concentrations, and that initiation with them stimulates promoter escape. Efficiency of initiation with NAD+, but not with UDP-containing factors, is affected by amino acids of the Rifampicin-binding pocket, suggesting altered RNA capping in Rifampicin-resistant strains. However, relative affinity to NAD+ does not depend on the -1 base of the template strand, as was suggested earlier. We show that incorporation of mature cell wall precursor, UDP-MurNAc-pentapeptide, is inhibited by region 3.2 of σ subunit, possibly preventing targeting of RNA to the membrane. Overall, our in vitro results propose a wide repertoire of potential bacterial RNA capping molecules, and provide mechanistic insights into their incorporation.
[Mh] Termos MeSH primário: Proteínas de Bactérias/genética
Parede Celular/genética
RNA Polimerases Dirigidas por DNA/genética
Capuzes de RNA/genética
Transcrição Genética
[Mh] Termos MeSH secundário: Proteínas de Bactérias/metabolismo
Sequência de Bases
Parede Celular/metabolismo
RNA Polimerases Dirigidas por DNA/metabolismo
Flavina-Adenina Dinucleotídeo/metabolismo
NAD/metabolismo
Capuzes de RNA/metabolismo
Homologia de Sequência do Ácido Nucleico
Uridina Difosfato Glucose/metabolismo
Uridina Difosfato N-Acetilglicosamina/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (RNA Caps); 0U46U6E8UK (NAD); 146-14-5 (Flavin-Adenine Dinucleotide); 528-04-1 (Uridine Diphosphate N-Acetylglucosamine); EC 2.7.7.6 (DNA-Directed RNA Polymerases); V50K1D7P4Y (Uridine Diphosphate Glucose)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171023
[Lr] Data última revisão:
171023
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170523
[St] Status:MEDLINE
[do] DOI:10.1093/nar/gkx452


  7 / 1046 MEDLINE  
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[PMID]:28423179
[Au] Autor:Tanaka Y; Ogawa T; Maruta T; Yoshida Y; Arakawa K; Ishikawa T
[Ad] Endereço:Department of Life Science and Biotechnology, Faculty of Life and Environmental Science, Shimane University, Matsue, Shimane, Japan.
[Ti] Título:Glucan synthase-like 2 is indispensable for paramylon synthesis in Euglena gracilis.
[So] Source:FEBS Lett;591(10):1360-1370, 2017 May.
[Is] ISSN:1873-3468
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:The phytoflagellate Euglena gracilis produces a large amount of paramylon (PM), a conglomerate of liner ß-1,3-glucan chains, as a storage polysaccharide. PM is synthesized from uridine diphosphate-glucose, but its mechanism of formation is largely unknown. Two enzymes, glucan synthase-like (EgGSL) 1 and EgGSL2 were previously identified as candidates for PM synthesis in a Euglena transcriptome analysis. Here, we performed a reverse genetic analysis on these enzymes. Knockdown of EgGSL2, but not EgGSL1, significantly inhibits PM accumulation in Euglena cells. Additionally, ß-1,3-glucan synthesis is detected in a PM-associated membrane fraction extracted from Euglena cells. Our findings indicate that EgGSL2 is the predominant enzyme for PM biosynthesis.
[Mh] Termos MeSH primário: Euglena gracilis/metabolismo
Glucanos/biossíntese
Glucosiltransferases/genética
[Mh] Termos MeSH secundário: Euglena gracilis/genética
Técnicas de Silenciamento de Genes
Glucosiltransferases/metabolismo
Filogenia
Proteínas de Protozoários/metabolismo
Uridina Difosfato Glucose/metabolismo
[Pt] Tipo de publicação:LETTER
[Nm] Nome de substância:
0 (Glucans); 0 (Protozoan Proteins); 51052-65-4 (paramylon); EC 2.4.1.- (Glucosyltransferases); V50K1D7P4Y (Uridine Diphosphate Glucose)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:170919
[Lr] Data última revisão:
170919
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170420
[St] Status:MEDLINE
[do] DOI:10.1002/1873-3468.12659


  8 / 1046 MEDLINE  
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[PMID]:28238427
[Au] Autor:Loughney JW; Lancaster C; Price CE; Hoang VM; Ha S; Rustandi RR
[Ad] Endereço:Biologics and Vaccines, Merck & Co., Inc., Kenilworth, NJ, USA. Electronic address: John.Loughney@merck.com.
[Ti] Título:Development of a non-radiolabeled glucosyltransferase activity assay for C. difficile toxin A and B using ultra performance liquid chromatography.
[So] Source:J Chromatogr A;1498:169-175, 2017 May 19.
[Is] ISSN:1873-3778
[Cp] País de publicação:Netherlands
[La] Idioma:eng
[Ab] Resumo:Clostridium difficile infection (CDI) is the leading cause of gastroenteritis-associated death in the United States. The major virulent factors of C. difficile are toxin A (TcdA) and toxin B (TcdB). Toxicity is mediated by the glucosyltransferase domains on TcdA and TcdB wherein a glucose is transferred from UDP-glucose to Ras homolog family member A (RhoA) receptor. This modification results in disruption of critical cell signaling events. Vaccination against these toxins is considered the best way to combat the CDI. In order to produce non-toxic TcdA and TcdB antigens, their glucosyltransferase domains were genetically mutated to inactivate the toxin activity. We have developed a reverse phase ultra performance liquid chromatographic (RP-UPLC) method to measure this glucosyltransferase activity by separating RhoA and glucosylated RhoA. Glucosylated RhoA and RhoA have a retention time (RT) of 31.25 and 31.95min. We determine for the first time the glucosyltransferase kinetics (K and k ) of both full length TcdA and TcdB to RhoA and demonstrate that the genetically mutated TcdA and TcdB show no glucosyltransferase activity. Furthermore, two-dimensional electron microscopy (2D EM) data demonstrates that the overall global structures of mutated toxins do not change compared to native toxins.
[Mh] Termos MeSH primário: Proteínas de Bactérias/metabolismo
Toxinas Bacterianas/metabolismo
Cromatografia Líquida de Alta Pressão
Clostridium difficile/metabolismo
Enterotoxinas/metabolismo
Glucosiltransferases/metabolismo
Proteína rhoA de Ligação ao GTP/análise
[Mh] Termos MeSH secundário: Glicosilação
Seres Humanos
Cinética
Espectrometria de Massas
Microscopia Eletrônica
Uridina Difosfato Glucose/metabolismo
Proteína rhoA de Ligação ao GTP/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (Bacterial Toxins); 0 (Enterotoxins); 0 (tcdA protein, Clostridium difficile); 0 (toxB protein, Clostridium difficile); EC 2.4.1.- (Glucosyltransferases); EC 3.6.5.2 (rhoA GTP-Binding Protein); V50K1D7P4Y (Uridine Diphosphate Glucose)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:170906
[Lr] Data última revisão:
170906
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170228
[St] Status:MEDLINE


  9 / 1046 MEDLINE  
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[PMID]:28104756
[Au] Autor:López-Gutiérrez B; Dinglasan RR; Izquierdo L
[Ad] Endereço:ISGlobal, Barcelona Ctr. Int. Health Res. (CRESIB), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain.
[Ti] Título:Sugar nucleotide quantification by liquid chromatography tandem mass spectrometry reveals a distinct profile in sexual stage parasites.
[So] Source:Biochem J;474(6):897-905, 2017 Mar 07.
[Is] ISSN:1470-8728
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:The obligate intracellular lifestyle of and the difficulties in obtaining sufficient amounts of biological material have hampered the study of specific metabolic pathways in the malaria parasite. Thus, for example, the pools of sugar nucleotides required to fuel glycosylation reactions have never been studied in-depth in well-synchronized asexual parasites or in other stages of its life cycle. These metabolites are of critical importance, especially considering the renewed interest in the presence of -, -, and other glycans in key parasite proteins. In this work, we adapted a liquid chromatography tandem mass spectrometry (LC-MS/MS) method based on the use of porous graphitic carbon (PGC) columns and MS-friendly solvents to quantify sugar nucleotides in the malaria parasite. We report the thorough quantification of the pools of these metabolites throughout the intraerythrocytic cycle of The sensitivity of the method enabled, for the first time, the targeted analysis of these glycosylation precursors in gametocytes, the parasite sexual stages that are transmissible to the mosquito vector.
[Mh] Termos MeSH primário: Guanosina Difosfato Fucose/metabolismo
Guanosina Difosfato Manose/metabolismo
Açúcares de Guanosina Difosfato/metabolismo
Plasmodium falciparum/metabolismo
Uridina Difosfato Galactose/metabolismo
Uridina Difosfato Glucose/metabolismo
Uridina Difosfato N-Acetilgalactosamina/metabolismo
[Mh] Termos MeSH secundário: Cromatografia Líquida
Eritrócitos/parasitologia
Gametogênese/fisiologia
Guanosina Difosfato Fucose/análise
Guanosina Difosfato Manose/análise
Açúcares de Guanosina Difosfato/análise
Seres Humanos
Estágios do Ciclo de Vida/fisiologia
Plasmodium falciparum/crescimento & desenvolvimento
Espectrometria de Massas em Tandem
Uridina Difosfato Galactose/análise
Uridina Difosfato Glucose/análise
Uridina Difosfato N-Acetilgalactosamina/análise
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Guanosine Diphosphate Sugars); 15839-70-0 (Guanosine Diphosphate Fucose); 2956-16-3 (Uridine Diphosphate Galactose); 3123-67-9 (Guanosine Diphosphate Mannose); 5750-57-2 (guanosine diphosphate glucose); 7277-98-7 (Uridine Diphosphate N-Acetylgalactosamine); V50K1D7P4Y (Uridine Diphosphate Glucose)
[Em] Mês de entrada:1706
[Cu] Atualização por classe:170615
[Lr] Data última revisão:
170615
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170121
[St] Status:MEDLINE
[do] DOI:10.1042/BCJ20161030


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[PMID]:27935293
[Au] Autor:Mahalingan KK; Baskaran S; DePaoli-Roach AA; Roach PJ; Hurley TD
[Ad] Endereço:Department of Biochemistry and Molecular Biology, Indiana University School of Medicine , Indianapolis, Indiana 46202, United States.
[Ti] Título:Redox Switch for the Inhibited State of Yeast Glycogen Synthase Mimics Regulation by Phosphorylation.
[So] Source:Biochemistry;56(1):179-188, 2017 Jan 10.
[Is] ISSN:1520-4995
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Glycogen synthase (GS) is the rate limiting enzyme in the synthesis of glycogen. Eukaryotic GS is negatively regulated by covalent phosphorylation and allosterically activated by glucose-6-phosphate (G-6-P). To gain structural insights into the inhibited state of the enzyme, we solved the crystal structure of yGsy2-R589A/R592A to a resolution of 3.3 Å. The double mutant has an activity ratio similar to the phosphorylated enzyme and also retains the ability to be activated by G-6-P. When compared to the 2.88 Å structure of the wild-type G-6-P activated enzyme, the crystal structure of the low-activity mutant showed that the N-terminal domain of the inhibited state is tightly held against the dimer-related interface thereby hindering acceptor access to the catalytic cleft. On the basis of these two structural observations, we developed a reversible redox regulatory feature in yeast GS by substituting cysteine residues for two highly conserved arginine residues. When oxidized, the cysteine mutant enzyme exhibits activity levels similar to the phosphorylated enzyme but cannot be activated by G-6-P. Upon reduction, the cysteine mutant enzyme regains normal activity levels and regulatory response to G-6-P activation.
[Mh] Termos MeSH primário: Glicogênio Sintase/genética
Mutação
Saccharomyces cerevisiae/genética
[Mh] Termos MeSH secundário: Cristalização
Cristalografia por Raios X
Cisteína/química
Cisteína/genética
Cisteína/metabolismo
Ativação Enzimática/efeitos dos fármacos
Ativação Enzimática/genética
Glucose-6-Fosfato/metabolismo
Glucose-6-Fosfato/farmacologia
Glicogênio/metabolismo
Glicogênio Sintase/química
Glicogênio Sintase/metabolismo
Cinética
Modelos Moleculares
Oxirredução
Fosforilação
Domínios Proteicos
Multimerização Proteica
Estrutura Secundária de Proteína
Saccharomyces cerevisiae/enzimologia
Proteínas de Saccharomyces cerevisiae/química
Proteínas de Saccharomyces cerevisiae/genética
Proteínas de Saccharomyces cerevisiae/metabolismo
Uridina Difosfato Glucose/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Saccharomyces cerevisiae Proteins); 56-73-5 (Glucose-6-Phosphate); 9005-79-2 (Glycogen); EC 2.4.1.11 (Glycogen Synthase); K848JZ4886 (Cysteine); V50K1D7P4Y (Uridine Diphosphate Glucose)
[Em] Mês de entrada:1705
[Cu] Atualização por classe:170721
[Lr] Data última revisão:
170721
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:161210
[St] Status:MEDLINE
[do] DOI:10.1021/acs.biochem.6b00884



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